title: Interleukin-8 delays spontaneous and tumor necrosis factor-alpha-mediated apoptosis of human neutrophils.
abstract: During inflammation, polymorphonuclear neutrophils (PMN) are exposed to and influenced by various cytokines, including the chemoattractant interleukin-8 (IL-8). We tested the hypothesis that IL-8 affects apoptosis in PMN. We investigated which IL-8 receptor (RI or RII) might be involved, as well as the role of Bcl-2. Human PMN were isolated and cultured up to 30 hours. Apoptosis was detected by UV and light microscopy, as well as by DNA-fragmentation assay, and quantitated by flow cytometry. Interleukin-8 significantly delayed spontaneous apoptosis at 10, 20, and 30 hours in a dose-dependent fashion. Polymorphonuclear neutrophil treatment with the highest concentration of IL-8 (100 nM) decreased the percentage of apoptotic cells from 2.1 +/- 1.5 to 0.8 +/- 0.2 after 10 hours, from 31 +/- 14 to 8 +/- 5 after 20 hours, and from 47 +/- 15 to 18 +/- 8 after 30 hours of incubation (P < 0.05 for all time points, N = 6). Interleukin-8 also inhibited TNF alpha-mediated PMN apoptosis. Incubation with 20 ng/ml TNF alpha resulted in 23 +/- 6% apoptotic cells at four hours, whereas pretreatment with IL-8 (50 nM) decreased this percentage to 11 +/- 3 (N = 5, P < 0.05). We next studied the role of both types of IL-8 receptors, RI and RII, by comparing the effect of IL-8 and the product of growth-related oncogene alpha (Gro alpha) on PMN cultured for 20 hours. Both IL-8 and Gro alpha attenuated apoptosis, although IL-8 was more effective than Gro alpha. Bcl-2 was detected by intracellular fluorescent antibody cell sorter analysis, Western blot, and reverse transcription-polymerase chain reaction (RT-PCR). Neither resting PMN nor IL-8-treated neutrophils expressed BCL-2 protein, which was readily detected in control cells. Furthermore, we could not detect BCL-2 gene expression by RT-PCR. We conclude that IL-8 prolongs the lifespan of human neutrophils in vitro by delaying apoptosis. This effect may be important for a controlled and effective inflammatory response. The delay in apoptosis can be mediated by the IL-8 RII, while RI may provide an added effect. The actions of IL-8 on apoptosis are Bcl-2 independent.
authors.affiliation: Franz-Volhard Clinic, Humboldt University of Berlin, Germany.
authors.forename: R
authors.initials: R
authors.lastname: Kettritz
authors.forename: M L
authors.initials: ML
authors.lastname: Gaido
authors.forename: H
authors.initials: H
authors.lastname: Haller
authors.forename: F C
authors.initials: FC
authors.lastname: Luft
authors.forename: C J
authors.initials: CJ
authors.lastname: Jennette
authors.forename: R J
authors.initials: RJ
authors.lastname: Falk
mesh_terms: D015703:Antigens, CD / Q000502:physiology; D017209:Apoptosis / Q000187:drug effects*; D019254:Genes, bcl-2; D006801:Humans; D016209:Interleukin-8 / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects* / Q000502:physiology; D018123:Receptors, Interleukin / Q000502:physiology; D023062:Receptors, Interleukin-8A; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
chemical_list: D015703:Antigens, CD; D016209:Interleukin-8; D018123:Receptors, Interleukin; D023062:Receptors, Interleukin-8A; D014409:Tumor Necrosis Factor-alpha